The renewal proposal is designed to continue studies of gonadotropin storage with the use of electron microscopic-immunocytochemical techniques. Our focus during the next 3 years will involve a morphometric-densitometric study of serially sectioned secretion granules in cells containing both follicle stimulating hormone (FSH) and luteinizing hormone (LH) intact and castrated male rat pituitaries and in gonadotropes in monolayer culture following exposure to gonadotropin releasing hormone (GnRH). The serial granules will be taken from the current population of electron photomicrographs of 2700 serially gonadotropes and corticotropes from intact, castrated and adrenalectomized male rats. Additional experimental groups proposed in this renewal will yield more serial cells for analysis of cell type and granule content. In addition, the study has been expanded to include in vivo and in vitro experiments designed to test the source and significance of the intracellular GnRH and adrenocorticotropin (ACTH)-like activity found in some gonadotropes. Castrated, adrenalectomized and stressed rats will be studied to determine the effect of the physiologically altered state on GnRH and ACTH storage patterns. In vitro experiments with pituitary cells in monolayer culture will determine if gonadotropes sequester, produce or secrete GnRH and ACTH. Stains for opioids (enkephalins, beta-endorphin) and 1-10 lipotropin (LPH) will be developed to determine if the ACTH-like reactivity in gonadotropes includes other peptides in the ACTH-LPH family. Finally, on serial ultrathin sections, the stains developed above will be applied to human pituitaries to determine if similar storage patterns exist in the primate.